The objective is to understand how viral gene expression is regulated in both adenovirus infected cells and in cells transformed by the virus. I will pursue my recent discovery (Ricciardi, Jones, Cepko, Sharp, Roberts, in preparation) that an acidic protein of 51K daltons is required for expression of adjacent viral genes in infected cells. One aim is to purify the 51K acidic protein and to produce monoclonal antibodies against it, in addition to other early proteins. I will study the distribution of the 51K protein in the cell following microinjection, and determine whether it is associated with specific macromolecules. Essential features of this regulatory protein will be investigated by the use of host range (hr) mutants of adenovirus 5 (Ad5) which are defective in the gene that encodes the 51K protein (unpublished). These Ad5 hr mutants will be sequenced to confirm preliminary findings that hrl has a nonsense mutation and hr3 and hr5 each have missense mutations in the 51K gene. I will complete a detailed genomic map of early adenovirus mRNAs and proteins using hybridization-selection (Ricciardi et al., PNAS 76, 4927, 1979) and RNA sizing (Miller et al., JMB, in press). The kinetics of expression of specific RNAs and proteins will be studied during early infection using these techniques. In transformed cells, I will ask if the 51K protein is required to induce expression of 58K and 15K tumor antigen genes as well as other integrated viral genes. I will study this regulation by constructing cell lines which contain integrated viral genes defective for the 51K viral protein and study gene expression in these cells by the combined techniques of microinjection, hybridization-selection and monoclonal antibodies. I will also construct cDNA clones to study sequence arrangements of viral transcripts in transformed cells and determine if some of the long transcripts in these cells (unpublished) are exclusively viral in origin, or if they contain host sequences.